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Dissertation zugänglich unter
URN: urn:nbn:de:gbv:18-74449
URL: http://ediss.sub.uni-hamburg.de/volltexte/2015/7444/

The Role of KAP1 Posttranslational Modifications during Infection with Human Adenovirus Type 5

Rolle von KAP1 Posttranslationalen Modifikationen im Verlauf der Infektion mit Humanem Adenovirus Typ 5

Bürck, Carolin

 Dokument 1.pdf (14.457 KB) 

Basisklassifikation: 42.32
Institut: Biologie
DDC-Sachgruppe: Biowissenschaften, Biologie
Dokumentart: Dissertation
Hauptberichter: Dobner, Thomas (Prof. Dr.)
Sprache: Englisch
Tag der mündlichen Prüfung: 10.07.2015
Erstellungsjahr: 2015
Publikationsdatum: 28.07.2015
Kurzfassung auf Englisch: Cell survival requires mechanisms to recognize and repair DNA damage. The mechanisms involved in DNA damage response (DDR) have been intensively studied and comprise ATM-, ATR- and DNA-PK-dependent pathways, which can in severe cases initiate apoptosis. Modulation of chromatin structure is a critical step for DNA repair proteins gaining access to the DNA. A recently identified modulator of chromatin structure is the Survival-time associated PHD protein in Ovarian Cancer 1 (SPOC1). SPOC1 complexes with the cellular co-repressor KRAB-associated Protein 1 (KAP1) as well as with Histone Methyltransferases (HMTs) and the Nucleosome Remodeling and Deacetylase Complex (NuRD), resulting in chromatin condensation and heterochromatin formation after the repair of DNA double strand breaks (DSBs). KAP1 function is known to be regulated via several posttranslational modifications (PTMs), such as phosphorylation and SUMOylation. In this context, KAP1 is phosphorylated and deSUMOylated upon DSBs, leading to the dissociation of the repressive components from the damaged sites, facilitating efficient DNA repair, thereby underlining the importance of these PTMs in enlarging protein functions.
Human adenovirus type 5 (HAd5) contains a linear double-stranded genome, which is internalized into the host cell nucleus for virus replication. The free viral linear genome activates host-cellular DDR mechanisms after entering the host cell, including ATM, ATR and DNA-PK pathways. For efficient virus replication HAd5 gene products of the early regions 1 and 4 (E1 and E4) counteract these effects by inactivation of DDR components. While some of the components are targeted for degradation by the E1B-55K/E4orf6-dependent E3 ubiquitin ligase complex, other DDR factors are relocalized and/or inactivated to ensure proper virus replication. Increasing evidence suggests that modulation of chromatin remodeling factors is a crucial step in virus replication. Recently, we reported that the cellular Daxx/ATRX chromatin remodeling complex negatively regulates HAd5 replication and that this host-cellular antiviral defense is counteracted by virus-mediated proteasomal degradation during HAd5 infection. Furthermore, we recently identified SPOC1 as a restriction factor and binding partner of the viral core protein pVII, which is relocalized to viral replication centers (VRCs) and subsequently targeted for proteasomal degradation.
This work demonstrates that the cellular co-repressor KAP1 interacts with E1B-55K via its C-terminus as well as with the viral DNA binding factor E2A/DBP. This study provides the first molecular evidence that KAP1 negatively regulates HAd5 productive infection. Although KAP1 is not degraded during HAd5 infection, its antiviral response is counteracted by induction of PTMs of the cellular factor. In this context, it was observed that KAP1 is phosphorylated in a dose-dependent manner and that its phosphorylation is counteracted by the viral phosphoprotein E1B-55K early in infection, suggesting the repression of KAP1-responsive pro-apoptotic genes. Additionally, KAP1 is deSUMOylated, known to result in dissociation of the repressive complex from the DNA, followed by DNA relaxation and transcriptional activation. Interestingly, reduction of KAP1 SUMO modification requires the presence of SUMOylated E1B-55K, indicating a tight interplay between PTMs of KAP1 and the viral factor. The fact that our group recently showed an interaction of SPOC1 with pVII suggests that the SPOC1/KAP1 complex functions to maintain the chromatinized state of the viral genome early in infection, whereas KAP1 phosphorylation and deSUMOylation of KAP1 results in the onset of viral DNA synthesis.
Consistent with the hypothesis of a flexible regulation of KAP1 co-repressor function by PTMs, this work reveals that viral proteins differentially regulate KAP1 SUMOylation status in transfection. Thereby, proteins associated with the incoming virus particle as well as immediate early proteins induce an increase of KAP1 SUMOylation, while in the presence of early and late proteins, KAP1 SUMO modification is reduced. Furthermore, this study provides evidence that KAP1 facilitates SUMOylation of several HAd5 proteins, indicating its involvement in the regulation of functional and/or localization changes of the respective viral factors.


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